This proposal is to study the mechanism of evolution of new beta-galactosidases in Escherichia coli. The main objectives are two-fold: (1) to determine what aspects of enzyme activity are altered early in the evolutionary process and which later, and to compare newly evolved enzymes and their regulation with highly adapted systems, specifically the lactase operon, and (2) to assess the predictability and reproducibility of the evolutionary process. Strains of E. coli will be evolved by artificial selection into forms capable of growth on lactase. These will be characterized physiologically (i.e. growth on lactase, inducibility) and biochemically (activity on ONPCi and lactase, thermostability, ion stimulation, etc.). The locus that evolves into the new beta-galactosidase, called ebg, will be analyzed genetically by means of mutation, genetic mapping and complementation. BIBLIOGRAPHIC REFERENCE: Hall, B. G., and D. L. Hartl. 1975. Regulation of newly evolved enzymes. II. The ebg repressor. Genetics 81:427-435.